Magnetic bead suspension reagent, method for purifying nucleic acid and method for sorting nucleic acid

Abstract

The present disclosure provides a magnetic bead suspension reagent, a method for purifying nucleic acid by using the magnetic bead suspension reagent, and a method for sorting nucleic acid by using the magnetic bead suspension reagent. The magnetic bead suspension reagent includes magnetic beads and a surfactant, the surfactant includes a nonionic surfactant and an anionic/cationic surfactant, and both the nonionic surfactant and the anionic/cationic surfactant are present in the magnetic bead suspension reagent.

1 . A magnetic bead suspension reagent comprising:
 magnetic beads; and   a surfactant,   wherein the surfactant comprises a nonionic surfactant and an anionic/cationic surfactant, and both the nonionic surfactant and the anionic/cationic surfactant are present in the magnetic bead suspension reagent.   
     
     
         2 . The magnetic bead suspension reagent according to  claim 1 ,
 wherein a side chain of the anionic/cationic surfactant comprises a carbon chain, and a length of the carbon chain is greater than or equal to 10 carbon atoms and less than or equal to 20 carbon atoms.   
     
     
         3 . The magnetic bead suspension reagent according to  claim 2 ,
 wherein the surfactant is selected from a group consisting of Tween 20, polyethylene glycol octyl phenyl ether, sodium dodecyl sulfate, octadecyl diester quaternary ammonium salt.   
     
     
         4 . The magnetic bead suspension reagent according to  claim 3 , wherein the surfactant comprises Tween 20, sodium dodecyl sulfate and octadecyl diester quaternary ammonium salt. 
     
     
         5 . The magnetic bead suspension reagent according to  claim 1 , further comprising:
 a dehydrating agent; and   an ionic salt,   wherein a volume of the surfactant accounts for 0.5%˜5% of a volume of the magnetic bead suspension reagent, a mass of the dehydrating agent accounts for 20%˜30% of a mass of the magnetic bead suspension reagent, and a molarity of the ionic salt in the magnetic bead suspension reagent is 1 mol/L˜5 mol/L.   
     
     
         6 . The magnetic bead suspension reagent according to  claim 5 ,
 wherein the volume of the surfactant accounts for 0.5% of the volume of the magnetic bead suspension reagent, the mass of the dehydrating agent accounts for 30% of the mass of the magnetic bead suspension reagent, and the molarity of the ionic salt in the magnetic bead suspension reagent is 1.5 mol/L.   
     
     
         7 . The magnetic bead suspension reagent according to  claim 5 ,
 wherein the dehydrating agent comprises polyethylene glycol with at least one relative molecular mass, and the relative molecular mass of the polyethylene glycol is in a range of 8000˜15000.   
     
     
         8 . The magnetic bead suspension reagent according to  claim 5 ,
 wherein the ionic salt is selected from a group consisting of NaCl and MgCl 2 .   
     
     
         9 . The magnetic bead suspension reagent according to  claim 1 , further comprising:
 a buffer solution,   wherein the buffer solution comprises tris(hydroxymethyl)aminomethane hydrochloride, and a molarity of the buffer solution is 0.1 mmol/L˜10 mmol/L, and a pH value of the buffer solution is 7.5˜8.5.   
     
     
         10 . The magnetic bead suspension reagent according to  claim 1 , further comprising:
 a stabilizing solution,   wherein the stabilizing solution comprises ethylene diamine tetraacetic acid, and a molarity of the stabilizing solution is 0.5 mmol/L˜20 mmol/L.   
     
     
         11 . The magnetic bead suspension reagent according to  claim 1 ,
 wherein each of the magnetic beads is a superparamagnetic magnetic bead, and the superparamagnetic magnetic bead has a three-layer structure, an innermost layer of the three-layer structure is polystyrene, an intermediate layer of the three-layer structure is Fe 3 O 4 , and an outermost layer of the three-layer structure is a modified layer with carboxyl and/or silanol functional groups.   
     
     
         12 . The magnetic bead suspension reagent according to  claim 11 ,
 wherein a ratio of a mass of the intermediate layer of the magnetic bead to a mass of the magnetic bead is greater than 70%, and a particle size of the magnetic bead is 50 nm˜4.5 and a concentration of the magnetic beads in the magnetic bead suspension reagent is 1˜10 mg/mL.   
     
     
         13 . A method for purifying nucleic acids, comprising:
 mixing the magnetic bead suspension reagent according to  claim 1  with a sample solution comprising nucleic acids in a reaction tube to form a first mixed solution;   placing the reaction tube accommodating the first mixed solution on a magnetic stand until the first mixed solution clear, and then removing a supernatant in the reaction tube;   adding an eluent into the reaction tube to form a second mixed solution, placing the reaction tube accommodating the second mixed solution on the magnetic stand until the second mixed solution clear; and   collecting a supernatant in the reaction tube.   
     
     
         14 . The method according to  claim 13 , after the step of removing a supernatant in the reaction tube and before the step of adding an eluent into the reaction tube to form a second mixed solution, further comprising performing the following step at least once:
 adding an ethanol solution into the remaining solution in the reaction tube to form a third mixed solution, letting the third mixed solution stand until the third mixed solution clear, and then removing the supernatant comprising the ethanol solution.   
     
     
         15 . The method according to  claim 13 , wherein the step of adding an eluent into the reaction tube to form a second mixed solution, placing the reaction tube accommodating the second mixed solution on the magnetic stand until the second mixed solution clear comprises:
 taking the reaction tube out from the magnetic stand, adding Te buffer solution into the reaction tube at a ratio of 2 times a volume of the sample solution to form the second mixed solution, and then placing the reaction tube accommodating the second mixed solution on the magnetic stand, letting the second mixed solution stand until the second mixed solution clear.   
     
     
         16 . A method for sorting nucleic acids, comprising:
 mixing the magnetic bead suspension reagent according to  claim 1  with a sample solution comprising nucleic acids in a first reaction tube to form a first mixed solution;   placing the first reaction tube accommodating the first mixed solution on a magnetic stand until the first mixed solution clear, and then transferring a supernatant in the first reaction tube into a second reaction tube;   mixing the magnetic bead suspension reagent according to  claim 1  with the supernatant in the second reaction tube to form a second mixed solution;   placing the second reaction tube accommodating the second mixed solution on the magnetic stand until the second mixed solution clear, and then removing a supernatant in the second reaction tube;   adding an eluent into the second reaction tube to form a third mixed solution, and placing the second reaction tube accommodating the third mixed solution on the magnetic stand until the third mixed solution clear; and   collecting a supernatant in the second reaction tube.   
     
     
         17 . The method according to  claim 16 , wherein the step of placing the first reaction tube accommodating the first mixed solution on a magnetic stand until the first mixed solution clear, and then transferring a supernatant in the first reaction tube into a second reaction tube comprises:
 placing the first reaction tube accommodating the first mixed solution on the magnetic stand and letting the first mixed solution stand until the first mixed solution clear, so that the magnetic beads move to a bottom of the first reaction tube and adsorb nucleic acids with a first length, and then transferring the supernatant comprising nucleic acids with a second length and nucleic acids with a third length in the first reaction tube into the second reaction tube,   wherein the first length is greater than the second length and the third length.   
     
     
         18 . The method according to  claim 17 , wherein the step of placing the second reaction tube accommodating the second mixed solution on the magnetic stand until the second mixed solution clear, and then removing a supernatant in the second reaction tube comprises:
 placing the second reaction tube accommodating the second mixed solution on the magnetic stand and letting the second mixed solution stand until the second mixed solution clear, so that the magnetic beads move to a bottom of the second reaction tube and adsorb the nucleic acids with the second length, and then removing the supernatant comprising the nucleic acids with the third length in the second reaction tube,   wherein the second length is greater than the third length.   
     
     
         19 . The method according to  claim 18 , wherein the step of collecting a supernatant in the second reaction tube comprises:
 collecting the supernatant in the second reaction tube to obtain the nucleic acids with the second length.   
     
     
         20 . The method according to  claim 16 , after the step of removing a supernatant in the second reaction tube and before the step of adding an eluent into the second reaction tube to form a third mixed solution, further comprising performing the following step at least once:
 adding an ethanol solution into the remaining solution in the second reaction tube to form a fourth mixed solution, letting the fourth mixed solution stand until the fourth mixed solution clear, and then removing the supernatant comprising the ethanol solution.   
     
     
         21 . The method according to  claim 16 , wherein the step of adding an eluent into the second reaction tube to form a third mixed solution, and placing the second reaction tube accommodating the third mixed solution on the magnetic stand until the third mixed solution clear comprises:
 taking the second reaction tube out from the magnetic stand, adding Te buffer solution into the second reaction tube at a ratio of 2 times a volume of the sample solution to form the third mixed solution, then placing the second reaction tube accommodating the third mixed solution on the magnetic stand and letting the third mixed solution stand until the third mixed solution clear.